This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. It is estimated that Enteroccus faecalis causes 300,000-500,000 cases of urinary tract infection (UTI) each year in the United States. As a nosocomial uropathogen, E. faecalis ranks second only to E. coli in the number of nosocomial UTI's. The mucosal epithelium is an important physical barrier to microbial pathogens and is also thought to play an active role in sensing and responding to the presence of foreign invaders. Very little is known about the nature of the host response to Gram-positive uropathogens, including E. faecalis. In order to establish infection in the host, pathogens must adapt strategies to overcome this innate defense system. We have previously shown that pathogenic-derived lineages of E. faecalis possess an antiphagocytic capsule that protects the organism in vitro against neutrophil-mediated opsonophagocytosis. Roughly 50% of E. faecalis isolates possess the genetic locus that encodes the capsule biosynthetic machinary. In addition, the presence of the capsule enhances the survival of E. faecalis strains at extraintestinal sites. What role the capsule plays in establishing infection at the mucosal epithelium in the urinary tract is one focus of the present study. To address this question, we will use a recently established murine ascending UTI model and compare a wild-type encapsulated strain with two isogenic capsule mutants that either result in structural alterations to the capsule (cpsF) or eliminate capsule production altogether (cpsC). The outcome measures will be assessed by quantifying the bacterial burden in the urine, bladder, and kidneys of infected animals. Bladder and kidney tissues will also be processed to examine the histopathology resulting from these infections. In addition, cytokine and chemokine profiles will be analyzed from the urine of infected animals using a Luminex system, and RT-PCR will be perfomed on RNA isolated from bladder and kidney urothelial cells as a complimentary approach to determine the cytokine/chemokine source within the urinary tract. In addition to capsule, other microbial factors that could potentially alter clearance by the innate defense system include two extracellular proteases (gelatinase and serine protease). Isogenic protease mutants will also be compared in the UTI model to determine the relative contribution of each protease to the severity of the infection.